Gastroesophageal reflux (GERD) is a chronic inflammatory disease affecting millions of Americans leading to the development of gastric/intestinal metaplasia (Barrett's esophagus), which is a precursor for adenocarcinoma (Ca). Although data from the stomach and colon suggest that colonizing bacteria are essential in inflammation-induced cancer development, little is known about esophageal bacterial flora. The long-term goal of this project is to define the role of bacteria in the progression of GERD to Barrett's esophagus and Ca. Our hypothesis is that bacterial flora exist in the normal esophagus, changing in the evolution of GERD into Barrett's esophagus and Ca. The specific aims are to: 1) define at a population level the bacterial flora in the normal esophagus and the esophagus with GERD-related diseases, 2) define at a species level the bacterial flora in the normal esophagus and GERD-related diseases, 3) determine host humoral immune responses to esophageal bacteria in patients with GERD-related diseases and controls. To define bacterial flora at a population level. The total number of bacteria per biopsy will be determined by quantitative real-time PCR using universal bacterial 16S rDNA primers with DNA from each biopsy. Specimens from disease groups will be compared for bacterial density (square mm mucosa). To define the flora at a species level, biopsies will be analyzed using sequence-based universal bacterial16S PCR and cultivation. PCR products will be cloned and sequenced, and species identification accomplished by comparing the sequences with kno\\0'D bacterial 16S sequences. Biopsies also will be cultured in anaerobic, aerobic, and microaerobic conditions, and colonies biochemically defined to a species level. Each disease group will be compared for the species identified and their prevalence. Cultivable whole cell bacterial antigens will be used in ELISA to determine whether the hosts recognize their presence. Serum antibodies also will be examined using immonoblots to identify disease-specific antigens. Significance: (i) Bacterial flora in the GI tract play important roles in pathologic conditions including inflammation and neoplasia. The esophagus is the only part of the GI tract where little is known about the bacterial flora, and the proposed study will assess the existence and complexity of the flora. (ii) While GERD initially results from chemical damage, bacterial overgrowth may promote intestinal metaplasia of the esophagus. The proposed study allows qualitative and quantitative examination of this hypothesis.